The insect-borne protozoa Leishmania causes death and morbidity in millions of humans living in and traveling to the developing world. Wars and economic globalization expose even more Americans to these infections. Despite the efforts of many scientists throughout the world, immunoprotection against Leishmania and other protozoa remains an elusive target, and there are still no effective vaccines approved for use against protozoan diseases. Over many millennia protozoan parasites have evolved complex life cycles that facilitate transmission and evasion of host immune systems. These adaptations have been well described in many cases, but generating protective immunity in humans has so far proven an insurmountable challenge. Many attempts have been made using killed, attenuated, or subunit vaccines without success. The ideal vaccine would maximize immune stimulation and safety while minimizing toxicity or the capacity to cause inadvertent disease for the widest range of vaccinees. We are pursuing an innovative approach to this problem with the potential to impact millions of people worldwide. Recently, we have generated proof-of-concept data to support a completely novel vaccine technology which accomplishes these goals and could be applied to many protozoan diseases. The strategy utilizes a novel type of chemical sterilization called S-59 that results in DNA crosslinks when exposed to UVA radiation. We and others have successfully used this technology to develop safe and effective recombinant bacterial vaccines already. Most recently, we have demonstrated that S-59/UVA treatment of Leishmania results in a Killed But Metabolically Active (KBMA) organism capable of protecting rodents from infection in a model of visceral leishmaniasis (Leishmania that infects the visceral organs). We have also demonstrated a powerful complementarity of this strategy with novel Toll-like receptor agonists that enhance immune responses against infectious agents when simply applied to the skin. In this proposal we will amplify these proof-of-concept studies and expand them to rodent models of cutaneous leishmaniasis to determine the potential of this technology against these devastating parasitic diseases. We will also determine the fundamental immune mechanisms associated with the protective immunity induced by this vaccine. The experiments outlined here will provide further proof of concept with the ultimate goal of developing novel prophylactic and therapeutic vaccines against leishmaniasis. This technology, if successful, may also apply to other parasitic protozoa infecting humans, such as malaria, Chagas'disease (American Trypanosomiasis), and Sleeping Sickness (African Trypanosomiasis). These studies could potentially lead to safe, whole cell vaccines against other intracellular organisms with complex life cycles, such as mycobacterial diseases (Tuberculosis, Leprosy, and Buruli ulcer) in which antibiotic resistance is an urgent global threat and vaccines may provide the best long term solution. The approach is innovative, but based on a strong, yet simple rationale. The impact of these vaccines on humanity would be enormous if they are successful. Additionally, the current studies are designed to answer fundamental questions about immunity to parasitic protozoan infections that will help advance vaccine discovery regardless of approach. PUBLIC HEALTH RELEVANCE: Leishmaniasis is a parasitic infection that causes death and illness in millions of humans living in and traveling to the developing world. Wars and economic globalization expose even more Americans to these infections. Despite the efforts of many scientists throughout the world, vaccine protection against Leishmania and other protozoa remains an elusive target, and there are still no effective vaccines approved for use against these diseases. We are pursuing an innovative approach to this problem by developing a completely novel vaccine technology against Leishmaniasis with the potential to impact millions of people worldwide. If effective, this technology could potentially be applied to other infectious agents such as malaria and tuberculosis in the future.